Cloning and expression heterologous alanine dehydrogenase genes: Investigation of reductive amination potential of L-alanine dehydrogenases for green synthesis of alanine derivatives

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dc.authoridAktaş, Fatih/0000-0002-2031-298Xen_US
dc.authoridbinay, baris/0000-0002-6190-6549en_US
dc.authorscopusid58063124700en_US
dc.authorscopusid6602703134en_US
dc.authorscopusid7003653102en_US
dc.authorscopusid16309105400en_US
dc.authorscopusid23471733900en_US
dc.authorwosidAktaş, Fatih/HIK-0830-2022en_US
dc.contributor.authorDemir, Garip
dc.contributor.authorValjakka, Jarkko
dc.contributor.authorTurunen, Ossi
dc.contributor.authorAktas, Fatih
dc.contributor.authorBinay, Baris
dc.date.accessioned2024-08-23T16:04:45Z
dc.date.available2024-08-23T16:04:45Z
dc.date.issued2024en_US
dc.departmentDüzce Üniversitesien_US
dc.description.abstractUnnatural amino acids (UAAs) offer significant promise in a wide range of applications, including drug discovery, the custom design of peptides and proteins, and their utility and use as markers for monitoring molecular interactions in biological research. The synthesis of UAAs presents a formidable challenge and can be classified into two primary categories: enzymatic and chemical synthesis. Notably, the enzymatic route, specifically asymmetric synthesis, emerges as a an attractive method for procuring enantiopure UAAs with high efficiency, owing to its streamlined and concise reaction mechanism. The current study investigated the reductive amination activity mechanisms of alanine dehydrogenase (L-AlaDH), sourced from a combination of newly and previously characterized microorganisms. Our principal aim was to evaluate the catalytic efficiency of these L-AlaDH enzymes concerning a range of specific ketoacids and pyruvate to ascertain their capability for facilitating the production of both natural and unnatural amino acids. After the characterization processes, mutation points for TtAlaDH were determined and as a result of the mutations, mutants that could use ketocaproate and ketovalerate more effectively than the wild type were obtained. Among the enzymes studied, MetAlaDH exhibited the highest specific activity against pyruvate, 173 U/mg, and a KM value of 1.3 mM. VlAlaDH displayed the most favourable catalytic efficiency with a rate constant of 170 s- 1mM- 1. On the other hand, AfAlaDH demonstrated the highest catalytic efficiency against alpha-ketobutyrate (34.0 s- 1mM-1) and alpha-ketovalerate (2.7 s- 1mM-1). Of the enzymes investigated in the study, TtAlaDH exhibited the highest effectiveness among bacterial enzymes in catalyzing ketocaproate with a measured catalytic efficiency of about 0.6 s- 1mM-1 and a KM value of approximately 0.3 mM. These findings provide valuable insights into the substrate specificity and catalytic performance of L-AlaDHs, enhancing our understanding of their potential applications in various biocatalytic processes.en_US
dc.description.sponsorshipScientific and Technological Research Council of Turkey (TUBITAK) [120Z501]en_US
dc.description.sponsorshipThis work was supported partially by The Scientific and Technological Research Council of Turkey (TUBITAK) (Project number: 120Z501) . We express our gratitude to Bauzyme for their valuable support. The authors would like to thank Professor of Organic Chemistry Janne Janis for his guidance in understanding the reaction mechanism.en_US
dc.identifier.doi10.1016/j.heliyon.2024.e26899
dc.identifier.issn2405-8440
dc.identifier.issue5en_US
dc.identifier.pmid38463761en_US
dc.identifier.scopus2-s2.0-85186655254en_US
dc.identifier.scopusqualityQ1en_US
dc.identifier.urihttps://doi.org/10.1016/j.heliyon.2024.e26899
dc.identifier.urihttps://hdl.handle.net/20.500.12684/14347
dc.identifier.volume10en_US
dc.identifier.wosWOS:001202266900001en_US
dc.identifier.wosqualityN/Aen_US
dc.indekslendigikaynakWeb of Scienceen_US
dc.indekslendigikaynakScopusen_US
dc.indekslendigikaynakPubMeden_US
dc.language.isoenen_US
dc.publisherCell Pressen_US
dc.relation.ispartofHeliyonen_US
dc.relation.publicationcategoryMakale - Uluslararası Hakemli Dergi - Kurum Öğretim Elemanıen_US
dc.rightsinfo:eu-repo/semantics/openAccessen_US
dc.subjectUnnatural amino acidsen_US
dc.subjectL-Alanine dehydrogenaseen_US
dc.subjectReductive amination mechanismen_US
dc.subjectActive site modellingen_US
dc.subjectGreen synthesisen_US
dc.subjectUnnatural Amino-Acidsen_US
dc.subjectMycobacterium-Tuberculosisen_US
dc.subjectPurificationen_US
dc.subjectCyanobacteriumen_US
dc.subjectProteinsen_US
dc.subjectAntigenen_US
dc.subjectEnzymesen_US
dc.subjectApoen_US
dc.titleCloning and expression heterologous alanine dehydrogenase genes: Investigation of reductive amination potential of L-alanine dehydrogenases for green synthesis of alanine derivativesen_US
dc.typeArticleen_US

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